Data_Sheet_1_Astrocytic Ephrin-B1 Controls Synapse Formation in the Hippocampus During Learning and Memory.docx

Astrocytes play a fundamental role in synapse formation, pruning, and plasticity, which are associated with learning and memory. However, the role of astrocytes in learning and memory is still largely unknown. Our previous study showed that astrocyte-specific ephrin-B1 knock-out (KO) enhanced but ephrin-B1 overexpression (OE) in hippocampal astrocytes impaired contextual memory recall following fear conditioning. The goal of this study was to understand the mechanism by which astrocytic ephrin-B1 influences learning; specifically, learning-induced remodeling of synapses and dendritic spines in CA1 hippocampus using fear-conditioning paradigm. While we found a higher dendritic spine density and clustering on c-Fos-positive (+) neurons activated during contextual memory recall in both wild-type (WT) and KO mice, overall spine density and mEPSC amplitude were increased in CA1 neurons of KO compared to WT. In contrast, ephrin-B1 OE in hippocampal astrocytes impaired dendritic spine formation and clustering, specifically on c-Fos(+) neurons, coinciding with an overall decrease in vGlut1/PSD95 co-localization. Although astrocytic ephrin-B1 influenced learning-induced spine formation, the changes in astrocytic ephrin-B1 levels did not affect spine enlargement as no genotype differences in spine volume were observed between trained WT, KO, and OE groups. Our results suggest that a reduced formation of new spines rather than spine maturation in activated CA1 hippocampal neurons is most likely responsible for impaired contextual learning in OE mice due to abundantly high ephrin-B1 levels in astrocytes. The ability of astrocytic ephrin-B1 to negatively influence new spine formation during learning can potentially regulate new synapse formation at specific dendritic domains and underlie memory encoding.

星形胶质细胞在突触形成、修剪和可塑性中发挥着基础作用，这些作用与学习和记忆密切相关。然而，星形胶质细胞在学习和记忆中的作用仍大部分不为人知。我们先前的研究表明，在胆碱能星形胶质细胞中特异性敲除 Ephrin-B1 可增强，而 Ephrin-B1 过表达则损害了恐惧条件化后的情境记忆回忆。本研究旨在揭示星形胶质细胞中 Ephrin-B1 影响学习的机制；具体而言，是通过恐惧条件化范式来探讨学习诱导的 CA1 海马体突触和树突棘的重塑。我们发现，在野生型（WT）和敲除（KO）小鼠中，在情境记忆回忆期间被激活的 c-Fos 阳性（+）神经元上均出现了更高的树突棘密度和聚集，与 KO 相比，WT 小鼠的 CA1 神经元整体树突棘密度和 mEPSC 幅度有所增加。相反，在海马星形胶质细胞中 Ephrin-B1 过表达损害了树突棘的形成和聚集，特别是在 c-Fos(+) 神经元上，这与 vGlut1/PSD95 共定位的整体减少相一致。尽管星形胶质细胞中的 Ephrin-B1 影响了学习诱导的树突棘形成，但星形胶质细胞中 Ephrin-B1 水平的变化并未影响树突棘的扩大，因为训练的 WT、KO 和 OE 组之间没有观察到树突体积的基因型差异。我们的结果表明，由于星形胶质细胞中 Ephrin-B1 水平异常升高，导致 OE 小鼠情境学习受损的主要原因可能是新树突棘形成减少而非树突棘的成熟。星形胶质细胞中 Ephrin-B1 在学习过程中负向影响新树突棘形成的能力，可能调节特定树突域中的新突触形成，并作为记忆编码的基础。